Fermentations have already been used for a very long time to preserve foodstuffs for example. For this, either microorganisms are used, for example in the production of wine and beer or in the production of yoghurt and kefir, or enzymes direct, for example when souring milk using rennet. Fermentations can proceed under both aerobic and anaerobic conditions. A fermentation under anaerobic conditions is also called anaerobic fermentation. Fermentations are also used in the industrial-scale production of chemicals, for example in the production of citric acid or in the production of medicinal products such as antibiotics or insulin. A large number of reactors and procedures have been developed for these industrial processes. Thus the processes can be carried out continuously or batch-wise. With the reactors, types have been developed in which the microorganisms or the enzyme are arranged stationary in a reactor, for example in a packed bed, over which the substrate is then passed in liquid form. However, processes in which the microorganisms or the enzyme are distributed homogenously in the substrate are also known. The microorganisms or the enzyme can be immobilized on a solid support which is distributed in the substrate. However, a support is not necessary in many reactions.
The organic substrates that are used in such fermentations can vary greatly. For example, an aqueous glucose solution can be used as a basis, which serves as substrate or energy source for the microorganism during production of the desired substance. The organic substrate can be homogeneous and present for example as a solution. With other technical fermentations very heterogeneous organic substrates are used. For example in biogas production substrates are used which can also comprise solid constituents, wherein these solid constituents can also be present in the form of relatively large pieces. Typical organic substrates as used in biogas production are for example animal manure which can also be mixed with litter, plant chaff or sewage sludge.
Various microorganisms, for example bacteria, fungi or also cell cultures, are used for fermentations. If a fermentation is carried out with the help of microorganisms, in one variant of the procedure firstly the microorganisms can colonize the organic substrate. For this, in a start-up phase of the bioreactor the substrate is firstly seeded with the relevant microorganism, which then multiplies, accompanied by reaction of the organic substrate, and colonizes the organic substrate.
However, microorganisms as well as reactions catalyzed by same are also known in which the substrate is not colonized by the microorganisms. With this method variant of a fermentation, the microorganisms float in a nutrient solution and absorb dissolved nutrients via the surface. The microorganisms can have previously secreted enzymes in order, for example, to decompose solid substrates into smaller water-soluble components.
If the process is to be carried out continuously, fresh organic substrate is channelled into the reaction chamber of the reactor during the fermentation, while a corresponding quantity of spent substrate is discharged from the reactor. Microorganisms are also removed from the reaction chamber with the discharged spent substrate. This loss of microorganisms must be compensated by a corresponding growth. In particular with slow-growing organisms the growth rate can be the limiting factor for the capacity of the reactor. Below a critical value at which the quantity of renewable microorganisms is greater than the quantity of microorganisms discharged with the spent substrate, there is always a sufficient quantity of microorganisms available to sustain the fermentation. Then, for example, the quantity of fed organic substrate can form the limiting factor. If the quantity of fed organic substrate is further increased, a critical value is reached at which the quantity of renewable microorganisms corresponds to the quantity of microorganisms discharged with the spent substrate. The fermentation in the reaction chamber then proceeds in a stable manner. With a further increase in the quantity of fed substrate the renewable quantity of microorganisms can no longer compensate the loss in microorganisms discharged with the spent substrate. The microorganisms content of the reactor then falls constantly with the result that the fermentation no longer proceeds in a stable manner. The performance of the reactor and thus the yield of product decrease with a further increase in the quantity of fed substrate until fermentation finally stops.
An example of a fermentation with very slow-growing microorganisms is biogas production. During the production of biogas, in the final step with the help of methanogenic bacteria, methane is produced from H2 and CO2 or from ethyl acetate or other low-molecular compounds such as methylamine. This reaction takes place under strict anaerobic and reductive conditions. For thermodynamic reasons, methanogenic bacteria can achieve only an extremely small energy gain per reacted substrate molecule. Long generation times are therefore a necessary and inevitable consequence. For this reason, the start-up phase of newly charged biogas reactors also lasts a long time. Once a biogas reactor has finally reached its operational state, the growth rate of the slowest-growing microorganisms determines the maximum possible throughput of organic substrate.
In order, for a given reactor, to increase the throughput rate and thus the capacity of the reactor, the stationary concentration of microorganisms in the reactor must be increased. This can take place within a very limited framework by optimizing process parameters. A further possibility for increasing the stationary quantity of microorganisms in the reaction chamber of the bioreactor is to recover the microorganisms from the discharged digestate and return them to the reaction chamber again. However, as the digestate is very heterogeneous in composition, the microorganisms can be separated from the digestate only with great difficulty.
A method is described in DE 10 2005 024 886 B3 in which magnetizable particles are added to the organic substrate. The microorganisms are present in the reaction chamber or in the organic substrate in the form of bacterial floc. The bacteria are surrounded by a layer of slime which makes possible a cohesion between the individual bacteria and the composition of larger aggregates. Magnetizable particles can be embedded in the slime separated from the microorganisms, whereby a force exerted on the magnetizable particles can be transferred onto the microorganisms. It was able to be shown that it is possible to separate the microorganisms from the digestate by passing the digestate past a stationary magnet. A separation tube, to the outside of which permanent magnets are attached, can be provided for this. If the digestate is passed through the separation tube the stationary magnet attracts the magnetizable particles enclosed in the bacteria slime, whereby the bacterial floc in the digestate is moved towards the magnet and deposited on the wall of the separation tube. If the permanent magnets are removed from the outer wall of the separation tube, the layer of microorganisms deposited on the inner wall of the separation tube can be flushed out of the tube. The collected microorganisms can then be returned to the reactor again.
Through the method described in DE 10 2005 024 886 B3 it was able to be shown that the separation of microorganisms with the help of magnetizable particles is possible. However, the method does not yet make possible such a high degree of separation that the method can be used profitably in industrial plants. The ferrite used in the method is used in very fine-particle form, with the result that its sedimentation rate is low and the particles can be enclosed in the layer of slime of the bacteria. These very small magnetizable particles move only very slowly in the applied magnetic field because of the viscosity of the water and the weak magnetic forces. If it is attempted to improve the separation rate of the magnetizable particles by increasing the force exerted by the magnets, the danger is that the magnetizable particles will be pulled out of the slime layer of the microorganism floc. It is then no longer possible to separate the bacterial floc from the digestate. If it is attempted to improve the separation rate by using larger magnetizable particles, these larger particles can be colonized by microorganisms, whereby the bonding of the microorganisms to the magnetizable particles improves. However, the problem then occurs that the sedimentation rate of the magnetizable particles is very high. The particles are then no longer distributed homogeneously in the organic substrate during the fermentation, i.e. the fermentation can be carried out uniformly in the whole reaction chamber only with considerable outlay.
The possibility of accumulating substances or microorganisms with the help of magnetizable particles from a mixture of substances has already been shown using various examples. However, these applications relate mostly only to the separation of very small quantities of substance, for example in use in test kits. The use of magnetizable particles then makes it possible to avoid a laborious separation by centrifuging.
Thus highly porous ferromagnetic or ferrimagnetic glass particles that contain iron oxide or iron-containing pigments are described in U.S. Pat. No. 8,202,427; U.S. Pat. No. 7,922,917; and, U.S. Pat. No. 7,183,002. The glass particles preferably have a diameter of 5 to 25 μm, particular preferably 7 to 10 μm. To produce the glass particles firstly a suspension of the magnetizable particles in for example glycerol or glycol is produced. A hydrolyzable silicon compound, for example a tetraalkoxy silane, is then added to this suspension. The silicon compound is then hydrolyzed with an alkaline or acidic buffer with the result that SiO2 deposits on the magnetizable particles and an open-pore structure forms. The particles are separated off and dried below the Curie temperature, preferably in the range of from 100 to 500° C. The glass particles can be used to accumulate a specific substance, for example a protein, from a sample mixture. For this the porous glass particle, the surface of which has optionally been modified by providing suitable groups in order to increase the affinity between the substance and the surface of the glass particle, is mixed with the sample mixture. The substance contained in the mixture is adsorbed on the surface of the magnetizable glass particles. By applying an external magnetic field the porous glass particles can be deposited together with the molecules of interest adsorbed on same for example on the wall of a sample vessel. The liquid phase can then be separated off very easily, with the result that only the magnetizable glass particles with the substance adsorbed thereon remain in the sample vessel.
M.-H. Liao, B.-H. Chen, Biotechnology Letters 24, 1913-1917, 2002 describe a magnetizable adsorbent which for example can be used to accumulate a protein from a solution. For this, nanoparticles from superparamagnetic Fe3O4 are coated with a covalently bonded layer of polyacrylic acid. The polyacrylate has a large number of ionic groups which can interact with the protein of interest, whereby the protein is bonded to the surface of the nanoparticle. The polyacrylate chains are bonded to the Fe3O4 nanoparticle by activation with carbodiimide. On average two polyacrylate molecules are bonded per magnetic nanoparticle.
X.-D. Tong, B. Xue, Y. Sun, Biotechnol. Prog. 2001, 17, 134-139 describe a magnetizable support which can be used for the adsorption of proteins as well as their accumulation. The magnetizable particles have a core of a magnetizable material, for example Fe3O4, which is coated with a shell of cross-linked polyvinyl alcohol. In order to increase the affinity of the particles for proteins, the surface can be modified by bonding corresponding groups to the surface of the particle.
In addition to the above-described magnetizable particles, a whole series of other supports are known for carrying out fermentations or accumulating proteins. Thus A. Soares, B. Guieysse, B. Mattiasson, Biotechnology Letters 25, 927-933, 2003 describe the biological degradation of nonylphenol in a continuously operated fixed bed bioreactor. The bed consists of a granular material of foam glass, wherein a biofilm of microorganisms is immobilized on the surface of the grains of the granular material. The substrate charged with nonylphenol is passed over the bed, wherein a clear degradation of the nonylphenol can be detected in the eluate removed from the reactor.
H. Zilouei, B. Guieysse, B. Mattiasson, Process Biochemistry 41 (2006), 1083-1089 describe the biological degradation of chlorophenols with the help of bacteria that are immobilized as biofilm in a packed bed of foam-glass granular material. Also here it was able to be shown that a clear degradation of the chlorophenols can be achieved if a substrate charged with chlorophenol is passed through the packed reactor.
Foam-glass granular materials are available on the market in various forms. The foam-glass granular materials can have an open-pore or a closed-pore structure. Moreover the granular materials can be produced from various types of glass. For example the strength and temperature resistance of the glass granular material can be increased by adding Al2O3.
The production of an open-pore foam-glass granular material is described in DE 195 31 801 A1. For this, a glass powder is mixed with a silicon organic compound as well as wax microspheres. This mixture is processed to a granular material which is firstly prehardened at a low temperature. A cohesion between the particles of the glass powder is thus produced by partial decomposition of the silicon organic compound. Then the wax is melted out by further increasing the temperature, with the result that open pores remain in the granular material. After the wax has been melted out the glass granular material is hardened at a higher temperature, preferably in the range of from 600 to 800° C. The open-pore glass granular material is suitable for immobilizing microorganisms which for example are used in aerobic waste-water treatment.
Likewise an open-pore foam-glass granular material which is produced from a glass powder mixture with a low-melting and a high-melting component is used in DE 197 34 791 A1. The glass powder mixture is mixed with a blowing agent and then heated above the blowing temperature of the low-melting glass component. The walls of the pores, which are constructed from the low-melting component, open during blowing, with the result that an open-pore structure is obtained.
This open-pore foam-glass granular material can also be used as a support for microorganisms for the aerobic treatment of waste waters as described in DE 198 17 268 A1. For this the open-pore foam-glass granular material is coated firstly with approximately 5 mass-% Fe2O3 by dipping the granular material into an iron salt solution and then tempering it. The granular material can then be fed into an activated sludge reactor. Through the iron oxide, poorly degradable compounds are pre-oxidized by the addition of hydrogen peroxide, while the further mineralization of microorganisms is effected.
Magnetizable foam-glass granular materials have also already been described. Thus foam glasses in which ferromagnetic substances, such as for example iron, nickel or cobalt, are homogenously distributed in the glass foam, are known from EP 1 900 698 A1 and EP 1 900 697 A1. By foam glass is meant a solidified glass foam which comprises air-tight closed cells. However, the production and the structure of the magnetizable glass foam are not described. The magnetizable foam glass is used to produce filters. These comprise a liquid-tight casing with foam glass shaped bodies contained therein. The shaped bodies can for example be a granular material with a predetermined or largely arbitrary granulation.
A further open-pore magnetizable inorganic material is described in U.S. Pat. No. 5,734,020. The porous glass is impregnated with a suspension of metallic particles to produce the material. Excess suspension is removed and the impregnated particles then dried. As the magnetizable particles are deposited in the pores of the glass foam, the pore volume of the glass decreases due to the impregnation. The granular material has a grain size in the range of from approximately 1 to 200 μm. It can be used as support for chromatographic processes, in immunoassays, for syntheses, for example of oligonucleotides, as well as other separating and purifying processes.
A granular material made of fragments of a sinter body sintered from ground blow-moulded glass in which iron particles are embedded is described in US2007/0104949. The sinter body is broken into a granular material with the result that a large surface area is obtained, wherein the iron particles protrude from the surface. The granular material is produced according to a specific embodiment by firstly mixing glass powder with a blowing agent and fine-particle iron and then blowing the mixture, wherein a body, for example in the form of a plate, is obtained. The plate is then broken down into a granular material. The granular material can be magnetically influenced with an iron content of 6 wt.-% and above. This can be used for example to separate off fine particles of the foam glass from a suspension of contaminants. The granular material can be used in particular as feedstock for water purification.
A method for producing foam-glass granular material which is suitable for example as additive for construction materials, to increase the insulating effect of for example walls, is described in US2007/0186587. To produce the foam-glass granular material, firstly pre-ground glass is mixed with water glass and blowing agent accompanied by the addition of water to form a crude mixture. The crude mixture is then wet-ground over several hours to form a slip. The slip is granulated to form granular material green bodies and these are foamed in rotary kilns at a temperature of usually 800 to 900° C.
A porous material which has a content of silicon oxide in a range of from 60 wt.-% to 85 wt.-%, aluminium oxide in a range of from 6 wt.-% to 20 wt.-% and alkaline- and/or alkaline-earth oxide and/or alkaline- and/or alkaline-earth hydroxide in a range of from 0 to 15 wt.-% is described in EP 2 022 768 A2. The surface of the porous material is surrounded by a skin which is preferably water-tight. The water absorptivity according to DIN EN 1609 is between 0 to 5 wt.-%, preferably 0.2 to 3 wt.-%, particularly preferably 0.5 to 1 wt.-%. The foam glass is produced by mixing clay, silicon oxide as well as optionally alkaline- and/or alkaline-earth oxide or alkaline- and/or alkaline-earth hydroxide with water to form a suspension. The suspension is granulated and the granular material optionally dried. The granular material is then heated, wherein a closed-pore porous material is obtained. In order to support the foaming, optionally a blowing agent can also be added to the suspension. Various fields of application are proposed for the foam-glass granular material, for example as thickener, thermal insulation material, sound-absorbing material, filler, construction material, fire-protection agent, refractory material, chromatography material and/or support material.
A further method for producing foam-glass granular material is described in US2008/0156038. Firstly a glass binder slop is produced from water, a propellant and a glass binder at room temperature. Glass powder is then added to the glass binder slop with the result that a moister, more stirrable glass mixture is obtained. The glass mixture is homogenized and then stirred for 2 to 6 hours to at least partly decompose the glass constituents. Further glass powder is then added and the mixture is granulated. The granular material is firstly dried and then foamed at a temperature of approximately 790° C.
As already explained, fermentations can be used for the production of a large number of compounds.
These can be relatively simple compounds such as methane but also compounds with very complex structure, such as for example in the production of medicinal products. The biocatalysts used in fermentations, such as enzymes or microorganisms, can be very valuable, whether they can be made available only at great cost or can be provided in a reactor only in a limited quantity, for example depending on the slow growth rate of a microorganism.
A further field in which great advantages can be expected if biologically active molecules that are accessible only with difficulty can be used repeatedly are separation processes. Capture molecules can be used to accumulate specific components from a solution. For example antibodies, DNA capture probes, RNA capture probes, protein A, avidin, streptavidin or proteins with long histidine chains can be used as capture molecules. Thus far the procedure has been to pass the solution that contains the molecules of interest over a column in which suitable capture molecules are immobilized. However, if the solution contains for example microorganisms, these must be separated off from the solution in advance.